Substituted 2-anilinopyrimidine derivatives as EGFR modulators

ABSTRACT

This application discloses novel substituted 2-anilinopyrimidine derivatives, and pharmaceutically acceptable salts, solvates, prodrugs, and compositions thereof, which are useful for the treatment or prevention of diseases or medical conditions mediated by epidermal growth factor receptors (EGFRs), including but not limited to a variety of cancers.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a Continuation of U.S. patent application Ser. No.15/534,838, filed Jun. 9, 2017, which is the U.S. National Phase ofInternational Patent Application No. PCT/US2015/065286, filed Dec. 11,2015, which claims priority under 35 U.S.C. § 119(e) to U.S. ProvisionalApplication No. 62/090,869, filed on Dec. 11, 2014, and No. 62/166,883,filed on May 27, 2015, all of which are incorporated herein by referencein their entireties.

FIELD OF THE INVENTION

The present invention relates to substituted 2-anilinopyrimidinederivatives and pharmaceutically acceptable salts and compositionsthereof useful for the treatment or prevention of diseases or medicalconditions mediated through mutated forms of epidermal growth factorreceptor (EGFR), such as various cancers.

BACKGROUND OF THE INVENTION

The epidermal growth factor receptor (EGFR, Her1, ErbB1) is a principalmember of the ErbB family of four structurally-related cell surfacereceptors with the other members being Her2 (Neu, ErbB2), Her3 (ErbB3)and Her4 (ErbB4). EGFR exerts its primary cellular functions though itsintrinsic catalytic tyrosine protein kinase activity. The receptor isactivated by binding with growth factor ligands, such as epidermalgrowth factor (EGF) and transforming growth factor-alpha (TGF-α), whichtransform the catalytically inactive EGFR monomer into catalyticallyactive homo- and hetero-dimers. These catalytically active dimers theninitiate intracellular tyrosine kinase activity, which leads to theautophosphorylation of specific EGFR tyrosine residues and elicits thedownstream activation of signaling proteins. Subsequently, the signalingproteins initiate multiple signal transduction cascades (MAPK, Akt andJNK), which ultimately mediate the essential biological processes ofcell growth, proliferation, motility and survival.

EGFR is found at abnormally high levels on the surface of many types ofcancer cells and increased levels of EGFR have been associated withadvanced disease, cancer spread and poor clinical prognosis. Mutationsin EGFR can lead to receptor overexpression, perpetual activation orsustained hyperactivity and result in uncontrolled cell growth, i.e.cancer. Consequently, EGFR mutations have been identified in severaltypes of malignant tumors, including metastatic lung, head and neck,colorectal and pancreatic cancers. In lung cancer, mutations mainlyoccur in exons 18 to 21, which encode the adenosine triphosphate(ATP)-binding pocket of the kinase domain. The most clinically relevantdrug-sensitive EGFR mutations are deletions in exon 19 that eliminate acommon amino acid motif (LREA) and point mutations in exon 21, whichlead to a substitution of arginine for leucine at position 858 (L858R).Together, these two mutations account for nearly 85% of the EGFRmutations observed in lung cancer. Both mutations have perpetualtyrosine kinase activity and as a result they are oncogenic. Biochemicalstudies have demonstrated that these mutated EGFRs bind preferentiallyto tyrosine kinase inhibitor drugs such as erlotinib and gefitinib overadenosine triphosphate (ATP).

Erlotinib and gefitinib are oral EGFR tyrosine kinase inhibitors thatare first line monotherapies for non-small cell lung cancer (NSCLC)patients having activating mutations in EGFR. Around 70% of thesepatients respond initially, but unfortunately they develop resistancewith a median time to progression of 10-16 months. In at least 50% ofthese initially responsive patients, disease progression is associatedwith the development of a secondary mutation, T790M in exon 20 of EGFR(referred to as the gatekeeper mutation). The additional T790M mutationincreases the affinity of the EGFR kinase domain for ATP, therebyreducing the inhibitory activity of ATP-competitive inhibitors likegefitinib and erlotinib.

Recently, irreversible EGFR tyrosine kinase inhibitors have beendeveloped that effectively inhibit the kinase domain of the T790M doublemutant and therefore overcome the resistance observed with reversibleinhibitors in the clinic. These inhibitors possess reactiveelectrophilic functional groups that react with the nucleophilic thiolof an active-site cysteine. Highly selective irreversible inhibitors canbe achieved by exploiting the inherent non-covalent selectivity of agiven scaffold along with the location of a particular cysteine residuewithin the ATP binding site. The acrylamide moieties of these inhibitorsboth undergo a Michael reaction with Cys797 in the ATP binding site ofEGFR^(T79OM) to form a covalent bond. This covalent mechanism is thoughtto overcome the increase in ATP affinity of the T790M EGRF double mutantand give rise to effective inhibition. However, these inhibitors maycause various undesired toxicities. Therefore, development of newinhibitors for treatment of various EGFR-related cancers is still inhigh demand.

SUMMARY OF THE INVENTION

The present invention provides novel compounds as EGFR tyrosine kinaseinhibitors that are therapeutically useful in the treatment orprevention of a number of EGFR-related diseases or disorders, such asvarious cancers.

In one aspect, the present invention provides compounds of formula I:

or a pharmaceutically acceptable salt, solvate, or prodrug thereof,wherein:

G is selected from substituted or unsubstituted 1H-indol-3-yl,substituted or unsubstituted 1H-indazol-3-yl, substituted orunsubstituted 2H-indazol-3-yl, and substituted or unsubstitutedpyrazolo[1,5-a]-pyridin-3-yl, and substituted or unsubstituted4,5,6,7-tetrahydropyrazolo[1,5-a]pyridin-3-yl;

X is selected from oxygen, sulfur, and methylene;

R¹ is selected from hydrogen, halogen, methyl, trifluoromethyl, andcyano;

R², R³, and R⁴ are the same or different and are independently selectedfrom hydrogen, halogen and trifluoromethyl;

R⁵ is selected from lower alkyl, optionally substituted 3- to 6-memberedheterocyclyl, R⁷R⁸N-(lower alkyl), and R⁷R⁸N-(cycloalkylalkyl), whereinR⁷ and R⁸ are the same or different and are independently selected fromhydrogen and lower alkyl; and

R⁶ is selected from lower alkoxy and lower alkyl.

In some preferred embodiments, in formula I, G is a 1H-indol-3-yl or1H-indazol-3-yl moiety having a formula

and the present invention provides a compound of formula II:

or a pharmaceutically acceptable salt, solvate, or prodrug thereof,wherein:

-   -   X is O, S, or CH₂;    -   Q is C—R¹⁰ or N    -   R⁹ is CH₃ or CH₂CH₂F; and    -   R¹⁰ is H or CH₃.

In some other preferred embodiments, in formula I, G ispyrazolo[1,5-a]-pyridin-3-yl, and the present invention provides acompound of formula V:

or a pharmaceutically acceptable salt, solvate, or prodrug thereof,wherein X is O, S, or CH₂.

In another aspect the present invention provides pharmaceuticalcompositions comprising any of the compounds, or a pharmaceuticallyacceptable salt, solvate, or prodrug thereof, and a pharmaceuticallyacceptable carrier.

The compounds and compositions of the present invention are useful fortreating diseases, disorders, or conditions associated with one or moreEGFR mutations. Such diseases, disorders, or conditions include thosedescribed herein, such as various cancers.

Thus, in another aspect, the present invention provides methods oftreating diseases or disorders associated with EGFR activities, such asvarious cancers associated with one or more EGFR mutations, or use ofthe compounds or compositions in the manufacture of medicaments fortreatment of these diseases or disorders.

In another aspect, the compounds of this invention are useful for thestudy of kinases in biological and pathological phenomena; the study oftransduction pathways mediated by such kinases; and the comparativeevaluation of new kinase inhibitors.

In another aspect, the present invention provides methods ofsynthesizing the compounds disclosed herein.

Other aspects or advantages of the present invention will be betterappreciated in view of the detailed description and claims.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 illustrates the H1975 tumor growth inhibition assay results forExample 1 in mice.

FIG. 2 illustrates the H1975 tumor growth inhibition assay results forExample 2 in mice.

FIG. 3 illustrates the HCC827 tumor growth inhibition assay results forExample 1 in mice.

FIG. 4 illustrates the average concentrations of Example 1 in plasma,brain and tumor tissues in mice following oral administration of a 25mg/kg dose in the HCC827 mouse xenograft model.

DETAILED DESCRIPTION OF THE INVENTION

In one aspect, the present invention provides a compound of the formulaI:

or a pharmaceutically acceptable salt, solvate, or prodrug thereof,wherein:

G is selected from the group consisting of substituted or unsubstituted1H-indol-3-yl, substituted or unsubstituted 1H-indazol-3-yl, substitutedor unsubstituted 2H-indazol-3-yl, substituted or unsubstitutedpyrazolo[1,5-a]-pyridin-3-yl, and substituted or unsubstituted4,5,6,7-tetrahydropyrazolo[1,5-a]pyridin-3-yl;

-   -   X is oxygen, sulfur, or methylene;    -   R¹ is hydrogen, halogen, methyl, trifluoromethyl, or cyano;

R², R³, and R⁴ are the same or different and are independently selectedfrom the group consisting of hydrogen, halogen, and trifluoromethyl;

R⁵ is selected from the group consisting of lower alkyl, optionallysubstituted 3- to 6-membered heterocyclyl, R⁷R⁸N-(lower alkyl), andR⁷R⁸N-(cycloalkylalkyl), wherein R⁷ and R⁸ are the same or different andare each independently selected from hydrogen and lower alkyl; and

R⁶ is lower alkoxy or lower alkyl.

In one embodiment of this aspect, G is selected from the groupconsisting of 1H-indol-3-yl, 1-methyl-1H-indol-3-yl,1-(2-fluoroethyl)-1H-indol-3-yl, 1,2-dimethyl-1H-indol-3-yl,pyrazolo[1,5-a]-pyridin-3-yl,4,5,6,7-tetrahydropyrazolo[1,5-a]pyridin-3-yl, 1-methyl-1H-indazol-3-yl,and 2-methyl-2H-indazol-3-yl.

In a preferred embodiment, G is selected from the group consisting of1-methyl-1H-indol-3-yl, 1-(2-fluoroethyl)-1H-indol-3-yl,1,2-dimethyl-1H-indol-3-yl, pyrazolo[1,5-a]-pyridin-3-yl, and1-methyl-1H-indazol-3-yl.

In a more preferred embodiment, G is 1-methyl-1H-indol-3-yl,1-(2-fluoroethyl)-1H-indol-3-yl, or 1,2-dimethyl-1H-indol-3-yl, and morepreferably 1-methyl-1H-indol-3-yl.

In another more preferred embodiment, G is pyrazolo[1,5-a]-pyridin-3-yl.

In another more preferred embodiment, G is 1-methyl-1H-indazol-3-yl.

In another embodiment of this aspect, R⁵ is selected from the groupconsisting of C₁-C₆ alkyl, substituted or unsubstituted azetidinyl,substituted or unsubstituted pyrrolidinyl, substituted or unsubstitutedpiperidinyl, R⁷R⁸N—(CH₂)_(n)— (n is an integer selected from 1 to 5),R⁷R⁸N—(C₃-C₆ cycloalkyl)-(CH₂)_(m)— (m=1, 2, 3), wherein R⁷ and R⁸ arethe same or different and are independently selected from hydrogen andlower alkyl.

In a preferred embodiment of this aspect, R⁵ is selected from the groupconsisting of methyl, 1-(dimethylamino)-cyclopropylmethyl,3-(dimethylamino)cyclobutyl, 1-methylazetidin-3-yl,(R)-1-methylpyrrolidin-3-yl, (S)-1-methylpyrrolidin-3-yl, and1-methylpiperidin-4-yl, and 2-dimethylamino-ethyl.

In a more preferred embodiment, R⁵ is 2-dimethylamino-ethyl [i.e.,(CH₃)₂NCH₂CH₂—].

In another embodiment of this aspect, R¹ is hydrogen, halogen, ormethyl.

In a preferred embodiment of this aspect, R¹ is hydrogen.

In another embodiment of this aspect, R² is hydrogen or halogen, whereinhalogen is preferably F or Cl.

In another embodiment of this aspect, R³ is hydrogen, F, Cl, or —CF₃.

In another embodiment of this aspect, R⁴ is hydrogen.

In another embodiment of this aspect, R² is hydrogen, F, or Cl; R³ ishydrogen, F, Cl, or —CF₃; and R⁴ is hydrogen.

In a preferred embodiment of this aspect, R², R³, and R⁴ are allhydrogen.

In a preferred embodiment of this aspect, R⁶ is lower alkoxy, preferablymethoxy or ethoxy.

In a more preferred embodiment, R⁶ is methoxy.

In another embodiment of this aspect, sometimes preferred, X is oxygen.

In another embodiment of this aspect, sometime preferred, X is sulfur.

In another embodiment of this aspect, sometimes preferred, X is —CH₂—.

As would be understood by a person skilled in the art, any plausible andstructurally allowable combinations of all the embodiments or preferredembodiments disclosed herein are encompassed and hereby specificallyincluded in the present invention.

For example, in some embodiments of this aspect, G is selected from thegroup consisting of 1H-indol-3-yl, 1-methyl-1H-indol-3-yl,1-(2-fluoroethyl)-1H-indol-3-yl, 1,2-dimethyl-1H-indol-3-yl,pyrazolo[1,5-a]-pyridin-3-yl,4,5,6,7-tetrahydropyrazolo[1,5-a]pyridin-3-yl, 1-methyl-1H-indazol-3-yl,and 2-methyl-2H-indazol-3-yl;

X is selected from the group consisting of oxygen, sulfur, andmethylene;

R¹ is selected from the group consisting of hydrogen, halogen, methyl,trifluoromethyl, and cyano;

R², R³, and R⁴ are the same or different and are independently selectedfrom the group consisting of hydrogen, halogen, and trifluoromethyl;

R⁵ is selected from the group consisting of1-(dimethylamino)-cyclopropylmethyl, 3-(dimethylamino)cyclobutyl,1-methylazetidin-3-yl, (R)-1-methylpyrrolidin-3-yl,(S)-1-methylpyrrolidin-3-yl, and 1-methylpiperidin-4-yl, and2-dimethylamino-ethyl; and

R⁶ is lower alkoxy.

In some preferred embodiments, G is a 1H-indol-3-yl or 1H-indazol-3-ylmoiety having a formula

and the present invention provides a compound of formula II:

or a pharmaceutically acceptable salt, solvate, or prodrug thereof,wherein:

-   -   X is O, S, or CH₂;    -   Q is C—R¹⁰ or N    -   R⁹ is CH₃ or CH₂CH₂F; and    -   R¹⁰ is H or CH₃.

In one preferred embodiment, in formula II, Q is C—R¹⁰, and the presentinvention provides a compound of formula III:

or a pharmaceutically acceptable salt, solvate, or prodrug thereof,wherein R⁹ is CH₃ or CH₂CH₂F; and R¹⁰ is H or CH₃.

In another preferred embodiment, in the compound of formula III, R⁹ isCH₃ and R¹⁰ is H.

In another preferred embodiment, in the compound of formula III, R⁹ isCH₃ and R¹⁰ is CH₃.

In another preferred embodiment, in the compound of formula III, R⁹ is2-fluoroethyl (FCH₂CH₂—), and R¹⁰ is H.

In another preferred embodiment, in formula III, R⁹ is CH₃, R¹⁰ is H,and X is O, the compound having the structure of formula 1:

In another preferred embodiment, in formula III, R⁹ is CH₃, R¹⁰ is CH₃,and X is O, the compound having the structure of formula 8:

In another preferred embodiment, in formula III, R⁹ is CH₃, R¹⁰ is H,and X is S, the compound having the structure of formula 2:

In another preferred embodiment, in formula III, R⁹ is CH₃, R¹⁰ is H,and X is CH₂, the compound having the structure of formula 4:

In another preferred embodiment, in formula III, R⁹ is —CH₂CH₂F, R¹⁰ isH, and X is O, the compound having the structure of formula 11:

In one preferred embodiment, in formula II, Q is N, and the presentinvention provides a compound of formula IV:

or a pharmaceutically acceptable salt, solvate, or prodrug thereof,wherein: R⁹ is H or CH₃; and X is O, S, or CH₂.

In a more preferred embodiment, in formula IV, R⁹ is H or CH₃, and X isO, the compound having the structure of formula 10:

In some other preferred embodiments, in formula I, G ispyrazolo[1,5-a]-pyridin-3-yl, and the present invention provides acompound of formula V:

or a pharmaceutically acceptable salt, solvate, or prodrug thereof,wherein X is O, S, or CH₂.

In a more preferred embodiment, in formula V, X is O, the compoundhaving the structure of formula 9:

In some other preferred embodiments, the present invention provides acompound selected from the group consisting of the Examples listed, or apharmaceutically acceptable salt, solvate, or prodrug thereof.

The more preferred compounds are listed below:

In another aspect, the present invention provides a pharmaceuticalcomposition comprising any one of the compounds of formulas I, II, III,IV, and V, or a pharmaceutically acceptable salt, solvate, or prodrugthereof, and a pharmaceutically acceptable carrier, adjuvant, diluent,and/or vehicle.

In one embodiment of this aspect, the composition further comprises asecond therapeutic agent.

In another embodiment of this aspect, the second therapeutic agent is adifferent EGFR modulator.

In another embodiment of this aspect, the second therapeutic agent is achemotherapeutic agent.

In another aspect, the present invention provides a method of treating adisease or disorder associated with an EGFR activity, comprisingadministration of a therapeutically effective amount of a compoundaccording to any one of formulas I, II, III, IV, and V, or apharmaceutically acceptable salt, solvate, prodrug, or a pharmaceuticalcomposition thereof, to a patient in need of treatment.

In one embodiment of this aspect, the disease or disorder is associatedwith one or more mutants of EGFR.

In another embodiment of this aspect, the mutant or mutants of EGFR areselected from L858R activating mutants L858R, delE746-A750, G719S; theExon 19 deletion activating mutant; and the T790M resistance mutant.

In another embodiment of this aspect, the disease or disorder is acancer.

In another embodiment of this aspect, the cancer is selected from braincancer, lung cancer, kidney cancer, bone cancer, liver cancer, bladdercancer, head and neck cancer, esophageal cancer, stomach cancer, coloncancer, rectum cancer, breast cancer, ovarian cancer, melanoma, skincancer, adrenal cancer, cervical cancer, lymphoma, and thyroid tumorsand their complications.

In another embodiment of this aspect, the method is used in conjunctionwith administering to the patient a second therapeutic agent.

In another embodiment of this aspect, the second therapeutic agent is achemotherapeutic agent.

In another embodiment of this aspect, the second therapeutic agent is adifferent EGFR modulator.

In another aspect, the present invention provides a method of inhibitinga mutant of EGFR in a subject, comprising contacting a biological sampleof said subject with a compound of any one of formulas I, II, III, IV,and V according to any embodiment disclosed herein, or apharmaceutically acceptable salt, solvate, or prodrug thereof. Suchinhibition can be in vitro or in vivo. If in vivo, the method maycomprise administering to said subject a compound of any one of formulasI, II, III, IV, and V according to any embodiment disclosed herein, or apharmaceutically acceptable salt, solvate, or prodrug thereof. If invitro, such inhibition may be conducted in a medium in any containerknown to those skilled in the art.

In another aspect, the present invention provides use of a compound ofany one of formulas I, II, III, IV, and V according to any embodimentdisclosed herein, or a pharmaceutically acceptable salt, solvate, orprodrug thereof, or a composition of any one of formulas I, II, III, IV,and V according to any embodiment disclosed herein, in the manufactureof a medicament for treatment of a disease or disorder associated withan EGFR activity.

In one embodiment of this aspect, disease or disorder is a cancerselected from the group consisting of brain cancer, lung cancer, kidneycancer, bone cancer, liver cancer, bladder cancer, head and neck cancer,esophageal cancer, stomach cancer, colon cancer, rectum cancer, breastcancer, ovarian cancer, melanoma, skin cancer, adrenal cancer, cervicalcancer, lymphoma, and thyroid tumors and their complications. In apreferred embodiment, the cancer is brain cancer or lung cancer. Thelung cancer includes, but is not limited to, non-small cell lung cancerand small cell lung cancer.

The terms in the present application, if not specifically defined, taketheir ordinary meanings as would be understood by those skilled in theart.

As used herein, the term “halo” or “halogen” refers to F, Cl, or Br.

The term “lower alkyl” refers to a branched or straight-chain alkylgroup having from one to seven carbon atoms, preferably one to four, andmore preferably one to two carbon atoms.

The term “lower alkoxy” refers to an alkoxy group (—OR) having from oneto seven, preferably one to four, and more preferably one to two carbonatoms.

The term “cyano” refers to —CN.

The term “pharmaceutically acceptable,” as used herein, refers to thosecompounds, materials, compositions, and/or dosage forms which are,within the scope of sound medical judgment, suitable for use in contactwith the tissues of patients without excessive toxicity, irritation,allergic response, or other problem or complication commensurate with areasonable benefit/risk ratio, and are effective for their intended use.

As used herein, the term “pharmaceutically acceptable salt” refers tothose salts which are, within the scope of sound medical judgment,suitable for use in contact with the tissues of humans and lower animalswithout undue toxicity, irritation, allergic response and the like, andare commensurate with a reasonable benefit/risk ratio. Pharmaceuticallyacceptable salts of the compounds of this invention include thosederived from suitable inorganic and organic acids and bases. Examples ofpharmaceutically acceptable, nontoxic acid addition salts are salts ofan amino group formed with inorganic acids such as hydrochloric acid,hydrobromic acid, phosphoric acid, sulfuric acid and perchloric acid orwith organic acids such as acetic acid, oxalic acid, maleic acid,tartaric acid, citric acid, succinic acid or malonic acid or by usingother methods used in the art such as ion exchange. Otherpharmaceutically acceptable salts include adipate, alginate, ascorbate,aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate,camphorate, camphorsulfonate, citrate, cyclopentanepropionate,digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate,glucoheptonate, glycerophosphate, gluconate, hemisulfate, heptanoate,hexanoate, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate,lactate, laurate, lauryl sulfate, malate, maleate, malonate,methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate,oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate,phosphate, pivalate, propionate, stearate, succinate, sulfate, tartrate,thiocyanate, p-toluenesulfonate, undecanoate, valerate salts, and thelike.

The term “solvate,” as used herein, means a physical association of acompound of this invention with a stoichiometric or non-stoichiometricamount of solvent molecules. For example, one molecule of the compoundassociates with one or more, preferably one to three, solvent molecules.It is also possible that multiple (e.g., two) molecules of the compoundshare one solvent molecule. This physical association may includehydrogen bonding. In certain instances the solvates will be capable ofisolation as crystalline solid. The solvent molecules in the solvate maybe present in a regular arrangement and/or a non-ordered arrangement.Exemplary solvates include, but are not limited to, hydrates,ethanolates, methanolates, and isopropanolates. Methods of solvation aregenerally known in the art.

The term “prodrug,” as used herein, refers to a derivative of a compoundthat can be transformed in vivo to yield the parent compound, forexample, by hydrolysis in blood. Common examples include, but are notlimited to, ester and amide forms of an active carboxylic acid compound;or vice versa, an ester from of an active alcohol compound or an amideform of an active amine compound. Such amide or ester prodrug compoundsmay be prepared according to conventional methods as known in the art.For example, a prodrug of a compound of formula II of the presentinvention could be in the form of the following formula VI:

wherein R^(x) and R^(y) are independently H and —C(O)—R, wherein R isC₁-C₄ alkyl, preferably methyl or ethyl, and more preferably methyl.Other prodrugs of the present invention can be prepared similarly fromany of formulas I, II, III, IV, and V.

When it is possible that, for use in therapy, therapeutically effectiveamounts of a compound of the present invention, or pharmaceuticallyacceptable salts or solvates thereof, may be administered as the rawchemical, it is possible to present the active ingredient as apharmaceutical composition. Accordingly, the disclosure further providespharmaceutical compositions, which include any compounds of the presentinvention, or pharmaceutically acceptable salts or solvates thereof, andone or more, preferably one to three, pharmaceutically acceptablecarriers, diluents, or other excipients. The carrier(s), diluent(s), orother excipient(s) must be acceptable in the sense of being compatiblewith the other ingredients of the formulation and not deleterious to thesubject being treated.

Pharmaceutical formulations may be presented in unit dose formscontaining a predetermined amount of active ingredient per unit dose.Typically, the pharmaceutical compositions of this disclosure will beadministered from about 1 to about 5 times per day or alternatively, asa continuous infusion. Such administration can be used as a chronic oracute therapy. The amount of active ingredient that may be combined withthe carrier materials to produce a single dosage form will varydepending on the condition being treated, the severity of the condition,the time of administration, the route of administration, the rate ofexcretion of the compound employed, the duration of treatment, and theage, gender, weight, and condition of the patient. Preferred unit dosageformulations are those containing a daily dose or sub-dose, as hereinabove recited, or an appropriate fraction thereof, of an activeingredient. Generally, treatment is initiated with small dosagessubstantially less than the optimum dose of the compound. Thereafter,the dosage is increased by small increments until the optimum effectunder the circumstances is reached. In general, the compound is mostdesirably administered at a concentration level that will generallyafford effective results without causing substantial harmful ordeleterious side effects.

When the compositions of this disclosure comprise a combination of acompound of the present disclosure and one or more, preferably one ortwo, additional therapeutic or prophylactic agent, both the compound andthe additional agent are usually present at dosage levels of betweenabout 10 to 150%, and more preferably between about 10 and 80% of thedosage normally administered in a monotherapy regimen.

Pharmaceutical formulations may be adapted for administration by anyappropriate route, for example, by the oral (including buccal orsublingual), rectal, nasal, topical (including buccal, sublingual, ortransdermal), vaginal, or parenteral (including subcutaneous,intracutaneous, intramuscular, intra-articular, intrasynovial,intrasternal, intrathecal, intralesional, intravenous, or intradermalinjections or infusions) route. Such formulations may be prepared by anymethod known in the art of pharmacy, for example by bringing intoassociation the active ingredient with the carrier(s) or excipient(s).Oral administration or administration by injection is preferred.

Pharmaceutical formulations adapted for oral administration may bepresented as discrete units such as capsules or tablets; powders orgranules; solutions or suspensions in aqueous or non-aqueous liquids;edible foams or whips; or oil-in-water liquid emulsions or water-in-oilemulsions.

For instance, for oral administration in the form of a tablet orcapsule, the active drug component can be combined with an oral,non-toxic pharmaceutically acceptable inert carrier such as ethanol,glycerol, water, and the like. Powders are prepared by comminuting thecompound to a suitable fine size and mixing with a similarly comminutedpharmaceutical carrier such as an edible carbohydrate, as, for example,starch or mannitol. Flavoring, preservative, dispersing, and coloringagent can also be present.

Capsules are made by preparing a powder mixture, as described above, andfilling formed gelatin sheaths. Glidants and lubricants such ascolloidal silica, talc, magnesium stearate, calcium stearate, or solidpolyethylene glycol can be added to the powder mixture before thefilling operation. A disintegrating or solubilizing agent such asagar-agar, calcium carbonate, or sodium carbonate can also be added toimprove the availability of the medicament when the capsule is ingested.

Moreover, when desired or necessary, suitable binders, lubricants,disintegrating agents, and coloring agents can also be incorporated intothe mixture. Suitable binders include starch, gelatin, natural sugarssuch as glucose or beta-lactose, corn sweeteners, natural and syntheticgums such as acacia, tragacanth or sodium alginate,carboxymethylcellulose, polyethylene glycol, and the like. Lubricantsused in these dosage forms include sodium oleate, sodium chloride, andthe like. Disintegrators include, without limitation, starch, methylcellulose, agar, betonite, xanthan gum, and the like. Tablets areformulated, for example, by preparing a powder mixture, granulating orslugging, adding a lubricant and disintegrant, and pressing intotablets. A powder mixture is prepared by mixing the compound, suitablecomminuted, with a diluent or base as described above, and optionally,with a binder such as carboxymethylcellulose, an aliginate, gelating, orpolyvinyl pyrrolidone, a solution retardant such as paraffin, aresorption accelerator such as a quaternary salt and/or and absorptionagent such as betonite, kaolin, or dicalcium phosphate. The powdermixture can be granulated by wetting with a binder such as syrup, starchpaste, acadia mucilage, or solutions of cellulosic or polymericmaterials and forcing through a screen. As an alternative togranulating, the powder mixture can be run through the tablet machineand the result is imperfectly formed slugs broken into granules. Thegranules can be lubricated to prevent sticking to the tablet formingdies by means of the addition of stearic acid, a stearate salt, talc, ormineral oil. The lubricated mixture is then compressed into tablets. Thecompounds of the present disclosure can also be combined with a freeflowing inert carrier and compressed into tablets directly without goingthrough the granulating or slugging steps. A clear or opaque protectivecoating consisting of a sealing coat of shellac, a coating of sugar orpolymeric material, and a polish coating of wax can be provided.Dyestuffs can be added to these coatings to distinguish different unitdosages.

Oral fluids such as solution, syrups, and elixirs can be prepared indosage unit form so that a given quantity contains a predeterminedamount of the compound. Syrups can be prepared by dissolving thecompound in a suitably flavored aqueous solution, while elixirs areprepared through the use of a non-toxic vehicle. Solubilizers andemulsifiers such as ethoxylated isostearyl alcohols and polyoxyethylenesorbitol ethers, preservatives, flavor additive such as peppermint oilor natural sweeteners, or saccharin or other artificial sweeteners, andthe like can also be added.

Where appropriate, dosage unit formulations for oral administration canbe microencapsulated. The formulation can also be prepared to prolong orsustain the release as for example by coating or embedding particulatematerial in polymers, wax, or the like.

It should be understood that in addition to the ingredients particularlymentioned above, the formulations may include other agents conventionalin the art having regard to the type of formulation in question, forexample those suitable for oral administration may include flavoringagents.

The term “patient” or “subject” includes both human and other mammals.

The term “mammal” or “mammalian animal” includes, but is not limited to,humans, dogs, cats, horses, pigs, cows, monkeys, rabbits and mice. Thepreferred mammals are humans.

The term “therapeutically effective amount” refers to an amount of acompound or composition that, when administered to a subject fortreating a disease, is sufficient to effect such treatment for thedisease. A “therapeutically effective amount” can vary depending on,inter alia, the compound, the disease and its severity, and the age,weight, or other factors of the subject to be treated. When applied toan individual active ingredient, administered alone, the term refers tothat ingredient alone. When applied to a combination, the term refers tocombined amounts of the active ingredients that result in thetherapeutic effect, whether administered in combination, serially, orsimultaneously.

The term “treating” or “treatment” refers to: (i) inhibiting thedisease, disorder, or condition, i.e., arresting its development; (ii)relieving the disease, disorder, or condition, i.e., causing regressionof the disease, disorder, and/or condition; or (iii) preventing adisease, disorder or condition from occurring in a subject that may bepredisposed to the disease, disorder, and/or condition but has not yetbeen diagnosed as having it. Thus, in one embodiment, “treating” or“treatment” refers to ameliorating a disease or disorder, which mayinclude ameliorating one or more physical parameters, though maybeindiscernible by the subject being treated. In another embodiment,“treating” or “treatment” includes modulating the disease or disorder,either physically (e.g., stabilization of a discernible symptom) orphysiologically (e.g., stabilization of a physical parameter) or both.In yet another embodiment, “treating” or “treatment” includes delayingthe onset of the disease or disorder.

When the term “about” is applied to a parameter, such as amount,temperature, time, or the like, it indicates that the parameter canusually vary by ±10%, preferably within ±5%, and more preferably within±2%. As would be understood by a person skilled in the art, when aparameter is not critical, a number provided in the Examples is oftengiven only for illustration purpose, instead of being limiting.

The term “a,” “an,” or “the,” as used herein, represents both singularand plural forms. In general, when either a singular or a plural form ofa noun is used, it denotes both singular and plural forms of the noun.

The following non-limiting Examples further illustrate certain aspectsof the present invention.

EXAMPLES

Chemical Synthesis

The compounds of the present invention are prepared generally accordingto Synthetic Schemes 1 to 8 in the illustrative, non-limiting Examplesdescribed below.

Abbreviations

The following abbreviations may be used:

-   THF=Tetrahydrofuran;-   conc.=concentrated-   DIEA=DIPEA=Diisopropylethylamine;-   sat.=saturated aqueous solution;-   FCC=flash column chromatography using silica;-   TFA=Trifluoroacetic acid;-   r.t.=room temperature;-   DI=deionized;-   DME=1,2-Dimethoxyethane-   DMF=N,N-Dimethylformamide;-   DMSO=Dimethylsulfoxide;-   DMA=N,N-Dimethylacetamide;-   HATU=O-(7-Azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluronium    hexafluoro-phosphate;-   EtOAc=Ethyl acetate;-   h=hour(s);-   NMM=N-Methylmorpholine-   Pd₂(dba)₃=Tris(dibenzylideneacetone)dipalladium(0);-   P(o-tol)₃=Tri(o-tolyl)phosphine.

Example 1N-(2-(2-(Dimethylamino)ethoxy)-4-methoxy-5-((4-(1-methyl-1H-indol-3-yl)pyrimidin-2-yl)amino)phenyl)acrylamide(1)

N-(4-(2-(Dimethylamino)ethoxy)-2-methoxy-5-nitrophenyl)-4-(1-methyl-1H-indol-3-yl)pyrimidin-2-amine(Scheme 1, Intermediate B). To a slurry of NaH (30 mmol, 60% oildispersion prewashed with hexanes) and 50 mL of 1,4-dioxane was added2-dimethylaminoethanol (27 mmol, 2.7 mL) dropwise with stirring underN₂. After stirring for 1 h, a slurry of A (5.4 mmol) in 50 mL of1,4-dioxane was added portion-wise over 15 min under a stream of N₂. Theresulting mixture was stirred overnight, then poured into water and thesolid was collected, rinsed with water, and dried under vacuum to yield2.6 g of product as a yellow solid. A purified sample was obtained fromchromatography (silica gel; CH₂Cl₂—CH₃OH gradient). ¹H NMR (300 MHz,DMSO) δ 2.26 (s, 6H), 2.70 (t, 2H, J=6 Hz), 3.87 (s, 3H), 4.01 (s, 3H),4.32 (t, 2H, J=6 Hz), 7.00-7.53 (m, 5H), 8.18-8.78 (m, 5H); C₂₄H₂₆N₆O₄m/z MH⁺ 463.

4-(2-(Dimethylamino)ethoxy)-6-methoxy-N1-(4-(1-methyl-1H-indol-yl)pyrimidin-2-yl)benzene-1,3-diamine(Scheme 1, Intermediate C). A suspension of 2.6 g of Intermediate B, 1.6g of Fe⁰, 30 mL of ethanol, 15 mL of water, and 20 mL of conc. HCl washeated to 78° C. for 3 h. The solution was cooled to room temperature,adjusted to pH 10 with 10% NaOH (aq) and diluted with CH₂Cl₂. Themixture was filtered through Dicalite, and the filtrate layers wereseparated. The aqueous phase was extracted with CH₂Cl₂ twice, and thecombined organic extracts were dried over Na₂SO₄ and concentrated.Column chromatography (silica gel, CH₂Cl₂—MeOH gradient) afforded 1.2 gof Intermediate C as a solid. C₂₄H₂₈N₆O₂ m/z MH⁺ 433.

N-(2-(2-(Dimethylamino)ethoxy)-4-methoxy-5-((4-(1-methyl-1H-indol-3-yl)pyrimidin-2-yl)amino)phenyl)acrylamide(1). To a solution of Intermediate C (2.8 mmol) in 50 mL of THF and 10mL of water was added 3-chloropropionychloride (2.8 mmol) dropwise withstirring. After 5 h of stirring, NaOH (28 mmol) was added and themixture was heated at 65° C. for 18 h. After cooling to roomtemperature, THF was partially removed under reduced pressure, and themixture was extracted with CH₂Cl₂, dried over Na₂SO₄, and concentrated.Chromatography of the crude product (silica gel, CH₂Cl₂-MeOH) afforded0.583 g of Example 1 as a beige solid. ¹H NMR (300 MHz, DMSO) δ 2.28 (s,6H), 2.50-2.60 (m, 2H), 3.86 (s, 3H), 3.90 (s, 3H), 4.19 (t, 2H, J=5.5Hz), 5.73-5.77 (m, 1H), 6.21-6.27 (m, 1H), 6.44-6.50 (m, 1H), 6.95 (s,1H), 7.11-7.53 (overlapping m, 3H), 7.90 (s, 1H), 8.27-8.30 (overlappingm, 3H), 8.55 (s, 1H), 8.84 (s, 1H), 9.84 (s, 1H) ppm; C₂₇H₃₀N₆O₃ m/z MH⁺487.

Example 2N-(2-((2-(Dimethylamino)ethyl)thio)-4-methoxy-5-((4-(1-methyl-1H-indol-3-yl)pyrimidin-2-yl)amino)phenyl)acrylamide(2)

N-(4-((2-(Dimethylamino)ethyl)thio)-2-methoxy-5-nitrophenyl)-4-(1-methyl-1H-indol-3-yl)pyrimidin-2-amine(Scheme 2, Intermediate D). To a slurry of NaH (54 mmol, 60% oildispersion prewashed with hexanes) and 25 mL of DMF was added a slurryof 2-dimethylaminoethanethiol hydrochloride (27 mmol) in 25 mL of DMFunder a stream of N₂. After stirring for 45 min, a slurry of A (5.4mmol) in 25 mL of DMF was added portionwise over 15 min to the mixtureunder a stream N₂. The resulting mixture was stirred overnight, thenpoured into water and the solid was collected, rinsed repeatedly withwater, and dried under vacuum to yield 2.5 g of product as a yellowsolid. C₂₄H₂₆N₆O₃S m/z MH⁺ 479.

4-((2-(Dimethylamino)ethyl)thio)-6-methoxy-N¹-(4-(1-methyl-1H-indol-3-yl)pyrimidin-2-yl)benzene-1,3-diamine(Scheme 2, Intermediate E). A suspension of 2.5 g of Intermediate D, 3.0g of Fe⁰, 50 mL of ethanol, 20 mL of water, and 7 mL of conc. HCl washeated to 78° C. for 3 h. The solution was cooled to room temperature,adjusted to pH 10 with 10% NaOH (aq), and diluted with CH₂Cl₂. Themixture was filtered through Dicalite, and the filtrate layers wereseparated. The aqueous phase was extracted with CH₂Cl₂ twice, and thecombined organic extracts were dried over Na₂SO₄ and concentrated.Column chromatography (silica gel, CH₂Cl₂-MeOH gradient) afforded 1.2 gof Intermediate E as a solid. C₂₄H₂₈N₆OS m/z MH⁺ 449.

N-(2-((2-(Dimethylamino)ethyl)thio)-4-methoxy-5-((4-(1-methyl-1H-indol-3-yl)pyrimidin-2-yl)amino)phenyl)acrylamide(2). To a solution of Intermediate E (2.7 mmol) in 50 mL of THF and 10mL of water was added 3-chloropropionychloride (4.0 mmol) dropwise withstirring. After 2 h of stirring, NaOH (27 mmol) was added and themixture was heated at 65° C. for 18 h. After cooling to roomtemperature, THF was partially removed under reduced pressure, and themixture was extracted with CH₂Cl₂, dried over Na₂SO₄, and concentrated.Chromatography of the crude product (silica gel, CH₂Cl₂-MeOH—NH₄OHgradient) afforded 0.622 g of Example 2 as an off-white solid: ¹H NMR(300 MHz, DMSO) δ 2.19 (s, 6H), 2.34 (t, 2H, J=6.5 Hz), 2.98 (t, 2H,J=6.5 Hz), 3.91 (s, 3H), 3.93 (s, 3H), 5.50-6.57 (overlapping m, 3H),7.12-9.88 (overlapping m, 10H), 10.17 (s, 1H) ppm. C₂₇H₃₀N₆O₂S m/z MH⁺503.

Example 3

N-(2,4-Dimethoxy-5-((4-(1-methyl-1H-indol-yl)pyrimidin-2-yl)amino)-phenyl)acrylamide(3)

N-(2,4-Dimethoxy-5-nitrophenyl)-4-(1-methyl-1H-indol-3-yl)pyrimidin-2-amine(Scheme 3, Intermediate F). Sodium methoxide, 25 wt. % solution inmethanol (40 mL, 175 mmol), was slowly poured into a stirred, ambienttemperature, suspension ofN-(4-fluoro-2-methoxy-5-nitrophenyl)-4-(1-methyl-1H-indol-3-yl)pyrimidin-2-amine(Scheme 1, Intermediate A; 5.8 g, 14.7 mmol) in methanol (125 mL) andheated at reflux for 4 days under nitrogen blanket, during which timethe solid did not dissolve. The reaction was cooled, product precipitateisolated by filtration, washed with cold methanol, and dried to yield5.45 g ofN-(2,4-dimethoxy-5-nitrophenyl)-4-(1-methyl-1H-indol-3-yl)pyrimidin-2-amine(Intermediate F) as a yellow powder. C₂₁H₁₉N₅O₄ m/z MH⁺ 406.

4,6-Dimethoxy-N¹-(4-(1-methyl-1H-indol-3-yl)pyrimidin-2-yl)benzene-1,3-diamine(Scheme 3, Intermediate G). Stannous chloride dihydrate (8.9 g, 39.4mmol) was added to a stirred, ambient temperature suspension ofN-(2,4-dimethoxy-5-nitrophenyl)-4-(1-methyl-1H-indol-3-yl)pyrimidin-2-amine(Intermediate F; 3.2 g, 7.9 mmol) in ethyl acetate (200 mL) and heatedat reflux under nitrogen blanket for 3 h. The reaction was allowed tocool, then poured into a 5% (w/v) solution of sodium bicarbonate in DIwater (400 mL) and stirred for 1 h. The multiphase mixture was thenfiltered through tightly packed Celite, with ethyl acetate rinsing ofthe filter cake. The filtrate was transferred to a separatory funnel andthe liquid phases separated. The retained ethyl acetate solution ofproduct was washed with brine and dried over anhydrous calcium sulfate.Filtration and evaporation yielded 1.6 g of crude product. Purificationby gradient flash chromatography (SiO₂, 0 to 70% hexanes/ethyl acetateover 20 min.) provided 0.9 g of4,6-dimethoxy-N¹-(4-(1-methyl-1H-indol-3-yl)pyrimidin-2-yl)benzene-1,3-diamine(Intermediate G) as a yellow foam. C₂₁H₂₁N₅O₂ m/z MH⁺ 376.

N-(2,4-Dimethoxy-5-((4-(1-methyl-1H-indol-3-yl)pyrimidin-2-yl)amino)phenyl)acrylamide(3). 3-Chloropropanoyl chloride (90 μL, 0.92 mmol) was rapidly added bysyringe to a rapidly stirred, ambient temperature, nitrogen blanketedsolution of4,6-dimethoxy-N¹-(4-(1-methyl-1H-indol-3-yl)pyrimidin-2-yl)benzene-1,3-diamine(Intermediate G; 351 mg, 0.94 mmol) and N-methylmorpholine (0.11 mL, 1.0mmol) in ethyl acetate (9.4 mL), precipitate immediately formed, andreaction was allowed to proceed for 40 min., evaporated to dryness, anddissolved in 10% (v/v) DI water/tetrahydrofuran. Solid sodium hydroxide(3 g, 75 mmol) was added and the stirred mixture heated to 50° C. for 17h. The reaction solution was cooled, partitioned between brine and ethylacetate. The ethyl acetate phase was dried over anhydrous calciumsulfate, filtered, and then chilled in an ice bath with stirring whileslowly being diluted with hexanes to precipitate the product. Thismaterial was isolated by filtration and dried to provide 189 mg ofExample 3 as fine light-yellow powder. ¹H NMR (300 MHz, DMSO) δ 3.88 (s,6H), 3.90 (s, 3H), 5.70 (dd, 1H, J=10.15, 1.92 Hz), 6.22 (dd, 1H,J=16.95, 2.03 Hz), 6.70 (q, 1H, J=9.06 Hz), 6.85 (s, 1H), 7.11-7.17 (m,2H), 7.23 (t, 1H, J=6.96 Hz), 7.50 (d, 1H, J=8.23 Hz), 7.93 (s, 1H),8.28 (m, 2H), 8.47 (s, 1H), 8.67 (s, 1H), 9.38 (s, 1H) ppm. ¹³C NMR (75MHz, DMSO) δ 33.4, 56.5, 56.7, 97.3, 107.1, 110.8, 113.0, 118.5, 119.5,121.3, 121.5, 122.3, 122.5, 125.9, 126.4, 132.8, 133.8, 138.1, 147.3,148.3, 157.8, 160.8, 162.3, 163.5 ppm. C₂₄H₂₃N₅O₃ m/z MH⁺ 430.

Example 4N-(2-(3-(Dimethylamino)propyl)-4-methoxy-5-((4-(1-methyl-1H-indol-3-yl)pyrimidin-2-yl)amino)phenyl)acrylamide(4)

N-(4-(3-(Dimethylamino)prop-1-yn-1-yl)-2-methoxy-5-nitrophenyl)-4-(1-methyl-1H-indol-3-yl)pyrimidin-2-amine(Scheme 4, Intermediate H). A solution of 3-dimethylamino-1-propyne(1.37 mL, 12.7 mmol) in 1,4-dioxane (60 mL) was treated with 1 M lithiumbis(trimethylsilyl)amide (12.7 mL, 12.7 mmol) and stirred for 30 min atRT under a nitrogen atmosphere. The resulting reaction mixture, whichappeared as a white slurry, was treated withN-(4-fluoro-2-methoxy-5-nitrophenyl)-4-(1-methyl-1H-indol-3-yl)pyrimidin-2-amine(Intermediate A; 1.00 g, 2.54 mmol) in one portion and heated at 80° C.while vigorously stirring under nitrogen for 5 h. The reaction mixturewas cooled to RT, quenched by the addition of 10 mL of water andsubsequently concentrated in vacuo. The residue was partitioned betweenwater (100 mL) and CH₂Cl₂ (50 mL). The basic aqueous layer was extractedwith CH₂Cl₂ (2×50 mL) and the combined organic extracts were washed withbrine (2×50 mL), dried (Na₂SO₄), filtered and concentrated in vacuo tofurnish 1.0 g of the crude product as a dark reddish-brown solid. Thismaterial was purified by gradient flash chromatography on SiO₂ elutingwith 0 to 10% methanol (containing 2% NH₄OH) in CH₂Cl₂ over 60 min toafford 98 mg ofN-(4-(3-(dimethylamino)prop-1-yn-1-yl)-2-methoxy-5-nitrophenyl)-4-(1-methyl-1H-indol-3-yl)pyrimidin-2-amine(Intermediate H) as an orange solid. C₂₅H₂₄N₆O₃ m/z MH⁺ 457.

4-(3-(Dimethylamino)propyl)-6-methoxy-N¹-(4-(1-methyl-1H-indol-3-yl)pyrimidin-2-yl)benzene-1,3-diamine(Scheme 4, Intermediate I). 10% Pd/C (10 mg) was added under a nitrogenatmosphere to a solution ofN-(4-(3-(dimethylamino)prop-1-yn-1-yl)-2-methoxy-5-nitrophenyl)-4-(1-methyl-1H-indol-3-yl)pyrimidin-2-amine(Intermediate H; 50 mg, 0.109 mmol) in 10 mL of THF/methanol (1:1). Ahydrogen-filled balloon was connected to the reaction vessel and thereaction was stirred at RT under a hydrogen atmosphere for 6 h. Thereaction mixture was filtered through Celite 545 and concentrated invacuo to give 50 mg of crude product. This material was purified bygradient flash chromatography on SiO₂ eluting with 0 to 10% methanol(containing 2% NH₄OH) in CH₂Cl₂ over 50 min to afford 34 mg of4-(3-(dimethylamino)propyl)-6-methoxy-N¹-(4-(1-methyl-1H-indol-3-yl)pyrimidin-2-yl)benzene-1,3-diamine(Intermediate I) as a foam. C₂₅H₃₀N₆O m/z MH⁺ 431.

N-(2-(3-(Dimethylamino)propyl)-4-methoxy-5-((4-(1-methyl-1H-indol-3-yl)pyrimidin-2-yl)amino)phenyl)acrylamide(4). 3-Chloropropanoyl chloride (18.2 μL, 0.190 mmol) was rapidly addedto a solution of4-(3-(dimethylamino)propyl)-6-methoxy-N¹-(4-(1-methyl-1H-indol-3-yl)pyrimidin-2-yl)benzene-1,3-diamine(Intermediate I; 34 mg, 0.079 mmol) in 3.2 mL of THF/water (9:1) whilestirring under nitrogen at RT. After 3 h, 1M aq NaOH (0.79 mL, 0.79mmol) and the reaction mixture was heated at 65° C. for 17 h. Thereaction mixture was cooled to RT, diluted with water (15 mL) and theresulting light gray precipitate was isolated by filtration to give 31mg of crude product. This material was purified by gradient flashchromatography on SiO₂ eluting with 0 to 10% methanol (containing 2%NH₄OH) in CH₂Cl₂ over 35 min to afford 22 mg of Example 4 as anoff-white solid. ¹H NMR (300 MHz, CDCl₃) δ 1.81-1.92 (m, 2H), 2.16 (t,2H, J=5.9 Hz), 2.27 (s, 6H), 2.69 (t, 2H, J=6.3 Hz), 3.89 (s, 3H), 3.98(s, 3H), 5.71 (dd, 1H, J=10.1, 1.9 Hz), 6.25 (dd, 1H, J=16.9, 10.1 Hz),6.48 (dd, 1H, J=16.9, 1.9 Hz), 6.66 (s, 1H), 7.17 (d, 1H, J=5.3 Hz),7.22-7.43 (m, 3H), 7.72 (s, 1H), 8.05-8.12 (m, 1H), 8.37 (d, 1H, J=5.3Hz), 8.85 (s, 1H), 9.33 (s, 1H), 10.95, (br s, 1H); C₂₈H₃₂N₆O₂ m/z MH⁺485.

Examples 5, 6, and 7

N-(2-(2-(Dimethylamino)ethoxy)-4-methoxy-5-((4-(2-methyl-2H-indazol-3-yl)pyrimidin-2-yl)amino)phenyl)acrylamide(5)

N-(2-((2-(Dimethylamino)ethyl)thio)-4-methoxy-5-((4-(2-methyl-2H-indazol-3-yl)pyrimidin-2-yl)amino)phenyl)acrylamide(6)

N-(2,4-Dimethoxy-5-((4-(2-methyl-2H-indazol-3-yl)pyrimidin-2-yl)amino)phenyl)acrylamide(7)

The synthesis ofN-(4-fluoro-2-methoxy-5-nitrophenyl)-4-(2-methyl-2H-indazol-3-yl)pyrimidin-2-amine(Intermediate J) is shown above in Scheme 5. Examples 5, 6, and 7 areprepared as in Schemes 1, 2, and 3, respectively, by substitutingIntermediate J for Intermediate A in each of those schemes.

Example 8N-(5-((4-(1,2-Dimethyl-1H-indol-3-yl)pyrimidin-2-yl)amino)-2-(2-(dimethylamino)ethoxy)-4-methoxyphenyl)acrylamide(8)

3-(2-Chloropyrimidin-4-yl)-1,2-dimethyl-1H-indole (Scheme 6,Intermediate K). Ferric chloride (5.8 g, 34.7 mmol) was rapidly added toa degassed, clear yellow solution of 1,2-dimethyl-1H-indole (4.9 g, 33.8mmol) and 2,4-dichloropyrimidine (5.2 g, 33.9 mmol) dissolved inanhydrous 1,2-dimethoxyethane (100 mL) while stirring at the ambienttemperature. The resultant black, opaque solution was stirred at ambienttemperature for 3 h under dry nitrogen atmosphere, then slowly pouredinto rapidly stirred 5% (w/v) aqueous NaHCO₃ (400 mL). Crude product wasisolated by filtration, and washed with DI water on the filter. Theprecipitate was suspended in methanol (200 mL) and evaporated to drynessto remove excess water, then triturated in hot acetonitrile, allowed tocool, and filtered to isolate 6.2 g of3-(2-chloropyrimidin-4-yl)-1,2-dimethyl-1H-indole (Intermediate K) as abrown powder. ¹H NMR (300 MHz, DMSO) δ 2.77 (s, 3H), 3.79 (s, 3H), 7.23(quin, 2H, J=7.53 Hz), 7.57 (d, 1H, J=7.25 Hz), 7.72 (d, 1H, J=5.61 Hz),8.10 (d, 1H, J=7.46 Hz), 8.61 (d, 1H, J=5.43 Hz) ppm. ¹³C NMR (75 MHz,DMSO) δ 12.8, 30.3, 108.8, 110.8, 117.5, 120.0, 121.8, 122.5, 125.8,137.4, 142.6, 159.8, 160.4, 165.2 ppm. C₁₄H₁₂ClN₃ m/z MH⁺ 258.

4-(1,2-Dimethyl-1H-indol-3-yl)-N-(4-fluoro-2-methoxy-5-nitrophenyl)-pyrimidin-2-amine(Scheme 6, Intermediate L). Reagent grade 1,4-dioxane (57 mL) was addedto a mixture of 3-(2-chloropyrimidin-4-yl)-1,2-dimethyl-1H-indole (1.47g, 5.70 mmol), 4-fluoro-2-methoxy-5-nitroaniline (1.06 g, 5.69 mmol),and p-toluenesulfonic acid monohydrate (1.31 g, 6.89 mmol) contained ina 100 mL round bottom flask fitted with a reflux condenser andblanketing nitrogen inlet. The magnetically stirred suspension washeated to reflux under nitrogen blanket. While approaching refluxtemperature the suspended solid dissolved. Reflux was continuedovernight, then the reaction was cooled and poured into rapidly stirringDI water (250 mL) to precipitate the product. Crude product was isolatedby filtration, washed with water and recrystallized from boiling2-propanol to yield 2.06 g of4-(1,2-dimethyl-1H-indol-3-yl)-N-(4-fluoro-2-methoxy-5-nitrophenyl)-pyrimidin-2-amine(Intermediate L) as a fine yellow powder. ¹H NMR (300 MHz, DMSO) δ 2.71(s, 3H), 3.78 (s, 3H), 4.01 (s, 3H), 7.10-7.20 (m, 3H), 7.41 (d, 1H,J=13.4 Hz), 7.55 (d, 1H, J=7.99 Hz), 7.98 (d, 1H, J=7.90 Hz), 8.44 (d,1H, J=5.70 Hz), 8.83 (br s, 1H), 8.93 (d, 1H, J=8.38 Hz). C₂₁H₁₈FN₅O₃m/z MH⁺ 408.

4-(1,2-Dimethyl-1H-indol-3-yl)-N-(4-(2-(dimethylamino)ethoxy)-2-methoxy-5-nitrophenyl)pyrimidin-2-amine (Scheme 6, Intermediate M). 2-Dimethylaminoethanol(0.43 mL, 4.27 mmol) was added, by syringe over 5 min., to a stirredsuspension of 60 wt. % sodium (173 mg, 4.33 mmol) in anhydrous1,4-dioxane at the ambient temperature. Gas evolution was readilyobserved. After ten min., with no further observable gas evolution,4-(1,2-dimethyl-1H-indol-3-yl)-N-(4-fluoro-2-methoxy-5-nitrophenyl)-pyrimidin-2-amine(Intermediate J) (351 mg, 0.86 mmol) was added, neat, to the rapidlystirred pot as one bolus. The reaction suspension immediately changed toa turbid red-brown color. After 5 min. an aliquot of the reaction waswithdrawn, quenched into DI water, and extracted into ethyl acetate.Analysis of this extract by UHPLC-MS revealed the reaction to becomplete. The pot contents were then poured into a stirred solution ofammonium chloride (0.23 g, 4.30 mmol) in DI water (150 mL) toprecipitate the product. The yellow precipitate was isolated byfiltration, washed with DI water, and allowed to dry to afford 386 mg of4-(1,2-dimethyl-1H-indol-3-yl)-N-(4-(2-(dimethylamino)ethoxy)-2-methoxy-5-nitrophenyl)pyrimidin-2-amine (Intermediate M). C₂₅H₂₈N₆O₄ m/z MH⁺=477.

N¹-(4-(1,2-Dimethyl-1H-indol-3-yl)pyrimidin-2-yl)-4-(2-(dimethylamino)-ethoxy)-6-methoxybenzene-1,3-diamine(Scheme 6, Intermediate N). Stannous chloride dihydrate (1.73 g, 7.67mmol) was added to a stirred suspension of4-(1,2-dimethyl-1H-indol-3-yl)-N-(4-(2-(dimethyl-amino)ethoxy)-2-methoxy-5-nitrophenyl)pyrimidin-2-amine (Intermediate M; 386 mg, 0.81 mmol) in ethyl acetate(40 mL) at the ambient temperature, and the mixture was heated at refluxunder nitrogen blanket for 17 h. The reaction was allowed to cool, thenpoured into a 1% (w/v) solution of sodium hydroxide in DI water (200 mL)and stirred for 1 h. The multiphase mixture was filtered throughtightly-packed Celite, with ethyl acetate rinsing of the filter cake.The filtrate was transferred to a separatory funnel and the liquidphases were separated. The retained ethyl acetate solution of productwas washed with brine, dried over anhydrous calcium sulfate, filteredand evaporated to provide a brown solid foam which was purified bygradient flash chromatography (SiO₂, 2% NH₄OH in MeOH/ethyl acetate, 0to 20% over 40 min.) to provide 186 mg ofN¹-(4-(1,2-dimethyl-1H-indol-3-yl)pyrimidin-2-yl)-4-(2-(dimethylamino)-ethoxy)-6-methoxybenzene-1,3-diamine(Intermediate N) as yellow solid. ¹H NMR (300 MHz, DMSO) δ 2.34 (s, 6H),2.70 (t, 2H, J=6.90 Hz), 2.75 (s, 3H), 3.58 (br s, 2H), 3.74 (s, 3H),3.83 (s, 3H), 4.07 (t, 2H, J=5.34 Hz), 6.57 (s, 1H), 6.95 (d, 1H, J=5.19Hz), 7.17-7.27 (m, 2H), 7.32-7.35 (m, 1H), 7.55 (s, 1H), 8.09 (dd, 1H,J=6.96, 1.77 Hz), 8.18 (s, 1H), 8.38 (d, 1H, J=5.22 Hz) ppm. C₂₅H₃₀N₆O₂m/z MH⁺=447.

N-(5-((4-(1,2-Dimethyl-1H-indol-3-yl)pyrimidin-2-yl)amino)-2-(2-(dimethyl-amino)ethoxy)-4-methoxyphenyl)acrylamide(8).N¹-(4-(1,2-Dimethyl-1H-indol-3-yl)pyrimidin-2-yl)-4-(2-(dimethylamino)ethoxy)-6-methoxy-benzene-1,3-diamine(Scheme 6, Intermediate N) is converted into Example 8 by reaction with3-chloropropionychloride followed by treatment with NaOH by using theprocedures described in the preparation of Example 1.

Example 9N-(2-(2-(Dimethylamino)ethoxy)-4-methoxy-5-((4-(pyrazolo[1,5-a]pyridin-3-yl)pyrimidin-2-yl)amino)phenyl)acrylamide(9)

N-(4-(2-(Dimethylamino)ethoxy)-2-methoxy-5-nitrophenyl)-4-(pyrazolo[1,5-a]pyridin-3-yl)pyrimidin-2-amine (Scheme 7, Intermediate P). To a slurry of NaH (21mmol, 60% oil dispersion prewashed with hexanes) and 20 mL of1,4-dioxane was added 2-dimethylaminoethanol (20 mmol, 2.4 mL) dropwisewith stirring under N₂. After stirring for 45 min, a slurry of compoundO (7.9 mmol) was added portion-wise, with stirring and under a stream ofN₂. The resulting mixture was stirred overnight, then poured into waterand the solid was collected, rinsed with water, and dried under vacuumto yield 1.7 g of Intermediate P as a yellow solid, which was used inthe next step without further purification: C₂₂H₂₃N₇O₄ m/z MH⁺450.

4-(2-(Dimethylamino)ethoxy)-6-methoxy-N1-(4-(pyrazolo[1,5-a]pyridin-3-yl)benzene)-1,3-diamine(Scheme 9, Intermediate Q). A suspension of 0.7 g of Intermediate P, 0.9g of Fe⁰, 7 mL of ethanol, 3 mL of water, and 2 mL of glacial aceticacid was heated to 78° C. for 1 h. The solution was cooled to roomtemperature, filtered through Dicalite, adjusted to pH 10 with 1 N NaOH(aq) and diluted with CH₂Cl₂. The filtrate layers were separated, andthe aqueous phase was extracted with CH₂Cl₂ twice, and the combinedorganic extracts were dried over Na₂SO₄ and concentrated. Columnchromatography (silica gel, CH₂Cl₂-MeOH gradient) afforded 0.28 g ofIntermediate Q as tan solid. C₂₂H₂₅N₇O₂ m/z MH⁺ 420.

N-(2-(2-(Dimethylamino)ethoxy)-4-methoxy-5-(4-(pyrazolo[1,5-a]pyridin-3-yl)benzene)-acrylamide(9). To a solution of Intermediate Q (0.6 g, 1.4 mmol) in 10 mL of THFand 4 mL of water was added 3-chloropropionychloride (0.15 mL, 1.6 mmol)dropwise with stirring. After 22 h of stirring, NaOH (0.7 g, 17 mmol)was added and the mixture was heated at 65° C. for 5 h. After cooling toroom temperature, THF was removed under reduced pressure, and themixture was extracted with CH₂Cl₂, dried over Na₂SO₄, and concentrated.Chromatography of the crude product (silica gel, CH₂Cl₂-MeOH) afforded0.294 g of Example 9 as a beige solid. C₂₅H₂₇N₇O₃ m/z MH⁺ 474. ¹H NMR(300 MHz, DMSO) δ 2.28 (s, 6H), 2.61-2.62 (m, 2H), 3.82 (s, 3H),4.20-4.22 (m, 2H), 5.69-5.73 (m, 1H), 6.20-6.22 (m, 1H), 6.42-6.48 (m,1H), 6.90-7.11 (m, 2H), 7.15-7.40 (m, 2H), 8.10-8.59 (overlapping m,4H), 8.72-8.96 (m, 2H), 10.13 (s, 1H) ppm.

Example 10N-(2-(2-(Dimethylamino)ethoxy)-4-methoxy-5-((4-(1-methyl-1H-indazol-3-yl)pyrimidin-2-yl)amino)phenyl)acrylamide(10)

N-(4-Fluoro-2-methoxy-5-nitrophenyl)-4-(1-methyl-1H-indazol-3-yl)pyrimidin-2-amine(Scheme 8, Intermediate R). Into a 1000-mL 3-necked round-bottom flaskpurged and maintained with an inert atmosphere of nitrogen, was placed asolution of 1H-indazole (10 g, 84.65 mmol, 1.00 equiv) inN,N-dimethylformamide (500 mL), I₂ (21.5 g, 84.65 mmol, 1.00 equiv).This was followed by the addition of KOH (19 g, 338.62 mmol, 4.00 equiv)in several batches at 0° C. The resulting solution was stirred overnightat room temperature. The reaction was then quenched by the addition of200 mL of aqueous Na₂S₂O₃. The resulting solution was extracted with3×500 mL of ethyl acetate and the organic layers combined. The resultingmixture was washed with 3×500 mL of brine. The mixture was dried overanhydrous sodium sulfate and concentrated under vacuum. The resultingmixture was washed with 1×100 mL of hexane. This resulted in 14 g (68%)of 3-iodo-1H-indazole as a white solid.

Into a 500-mL 3-necked round-bottom flask purged and maintained with aninert atmosphere of nitrogen, was placed a solution of3-iodo-1H-indazole (14 g, 57.37 mmol, 1.00 equiv) in tetrahydrofuran(200 mL). This was followed by the addition of NaH (65%) (2.5 g, 1.20equiv) in several batches at 0° C. The mixture was stirred for 30 min at0° C. To this was added iodomethane (9.7 g, 68.34 mmol, 1.20 equiv)dropwise with stirring at 0° C. The resulting solution was stirred for 1h at room temperature. The reaction was then quenched by the addition of300 mL of water/ice. The resulting solution was extracted with 2×300 mLof ethyl acetate and the organic layers combined. The resulting mixturewas washed with 1×300 mL of brine. The mixture was dried over anhydroussodium sulfate and concentrated under vacuum. The residue was appliedonto a silica gel column with ethyl acetate/petroleum ether (1:5). Thisresulted in 8 g (54%) of 3-iodo-1-methyl-1H-indazole as a yellow solid

Into a 500-mL 3-necked round-bottom flask purged and maintained with aninert atmosphere of nitrogen, was placed a solution of3-iodo-1-methyl-1H-indazole (5 g, 19.38 mmol, 1.00 equiv) in 1,4-dioxane(200 mL), hexamethyldistannane (12 g, 36.63 mmol, 2.00 equiv),tetrakis(triphenylphosphane) palladium (2.2 g, 1.90 mmol, 0.10 equiv).The resulting solution was stirred for 6 h at 100° C. The reactionmixture was cooled to room temperature with a water/ice bath. Thereaction was then quenched by the addition of 30 mL of aqueous KF (1 N)dropwise with stirring. The resulting solution was stirred for 0.5 h atroom temperature. The resulting solution was diluted with 200 mL of H₂O.The resulting solution was extracted with 2×200 mL of ethyl acetate andthe organic layers combined. The resulting mixture was washed with 3×200mL of brine. The mixture was dried over anhydrous sodium sulfate andconcentrated under vacuum. The residue was applied onto a silica gelcolumn with ethyl acetate/petroleum ether (1:5). This resulted in 3.9 g(68%) of 1-methyl-3-(trimethylstannyl)-1H-indazole as yellow liquid.

Into a 250-mL 3-necked round-bottom flask purged and maintained with aninert atmosphere of nitrogen, was placed1-methyl-3-(trimethylstannyl)-1H-indazole (3.9 g, 13.22 mmol, 1.00equiv), 1,4-dioxane (100 mL), 2,4-dichloropyrimidine (2.0 g, 13.42 mmol,1.00 equiv), tetrakis(triphenylphosphane) palladium (1.5 g, 1.30 mmol,0.10 equiv). The resulting solution was stirred overnight at 105° C. Thereaction mixture was cooled to room temperature with a water/ice bath.The reaction was then quenched by the addition of 200 mL of water/ice.The solids were collected by filtration. The filter cake was washed with1×100 mL of Et₂O. This resulted in 2.1 g (65%) of3-(2-chloropyrimidin-4-yl)-1-methyl-1H-indazole as a light yellow solid.

Into a 250-mL 3-necked round-bottom flask, was placed3-(2-chloropyrimidin-4-yl)-1-methyl-1H-indazole (2.9 g, 11.85 mmol, 1.00equiv), 4-fluoro-2-methoxy-5-nitroaniline (2.2 g, 11.82 mmol, 1.00equiv), 2-propanol (80 mL), TsOH (2.4 g, 13.94 mmol, 1.20 equiv). Theresulting solution was stirred overnight at 80° C. The reaction mixturewas cooled to room temperature with a water/ice bath. The solids werecollected by filtration. The filter cake was washed with 100 mL ofCH₃CN. The solid was dried in an oven. This resulted in 1.06 g (23%) ofN-(4-fluoro-2-methoxy-5-nitrophenyl)-4-(1-methyl-1H-indazol-3-yl)pyrimidin-2-amine(Intermediate R) as a yellow solid. (ES, m/z): [M+H]⁺=395; ¹H-NMR (300MHz, DMSO-d₆,) δ 8.96 (br, 1H), 8.87-8.85 (d, J=8.4 Hz, 2H), 8.56-8.54(d, J=5.4 Hz, 1H), 8.49-8.46 (d, J=8.1 Hz, 1H), 7.77-.775 (d, J=8.4 Hz,1H), 7.58-7.57 (d, J=5.1 Hz, 1H), 7.52-7.47 (t, J=7.2 Hz, 1H), 7.44-7.40(d, J=13.5 Hz, 1H), 7.26-7.21 (t, J=7.5 Hz, 1H), .4.19 (s, 1H), 4.01 (s,1H) ppm.

N-(4-(2-(Dimethylamino)ethoxy)-2-methoxy-5-nitrophenyl)-4-(1-methyl-1H-indazol-3-yl)pyrimidine-2-amine(Scheme 8, Intermediate S). To a suspension of NaH (31 mg, 1.3 mmol) in10 mL of 1,4-dioxane was added 2-dimethylaminoethanol (0.16 mL, 1.3mmol) dropwise with stirring under N₂. After stirring for 1.5 h,Intermediate R (0.2 g, 0.51 mmol) was added portionwise. After 0.5 h,the reaction mixture was quenched with water and extracted with CH₂Cl₂.The organic phase was dried over Na₂SO₄, filtered and concentrated toyield 0.23 g ofN-(4-(2-(dimethylamino)ethoxy)-2-methoxy-5-nitrophenyl)-4-(1-methyl-1H-indazol-3-yl)pyrimidine-2-amine(Intermediate S): m/z MH⁺=464.

4-(2-(Dimethylamino)ethoxy)-6-methoxy-N¹-(4-(1-methyl-1H-indazol-3yl)pyrimidin-2-yl)benzene-1,3-diamine(Scheme 8, Intermediate T). A suspension ofN-(4-(2-(dimethylamino)ethoxy)-2-methoxy-5-nitrophenyl)-4-(1-methyl-1H-indazol-3-yl)pyrimidine-2-amine(0.23 g), 0.28 g of Fe⁰, 10 mL of 70% ethanol/H₂O, and 0.5 mL of aceticacid was heated at reflux with stirring for 2 h. The mixture was cooledto room temperature, then filtered. The filtrate was adjusted to pH 10,then extracted with CH₂Cl₂. The organic phases were combined, dried overNa₂SO₄, filtered and concentrated. The crude product was purified bychromatography (silica gel, CH₂Cl₂-1% NH₄OH/MeOH gradient) to afford4-(2-(dimethylamino)ethoxy)-6-methoxy-N¹-(4-(1-methyl-1H-indazol-3yl)pyrimidin-2-yl)benzene-1,3-diamine(Intermediate T) as an off-white solid: m/z MH⁺ 434.

N-(2-(2-(Dimethylamino)ethoxy)-4-methoxy-5-((4-(1-methyl-1H-indazol-3-yl)pyrimidin-2-yl)amino)phenyl)acrylamide(Example 10). To a solution of4-(2-(dimethylamino)ethoxy)-6-methoxy-N1-(4-(1-methyl-1H-indazol-3yl)pyrimidin-2-yl)benzene-1,3-diamine(60 mg, 0.14 mmol) dissolved in 10 mL of 4:1 THF:H₂O was added3-chloropropionyl chloride (17 mg, 0.14 mmol). After 4 h, NaOH (1.4mmol, 56 mg) was added and the mixture was heated at reflux for 5 h. THFwas removed under reduced pressure, and the aqueous phase was extractedwith ethyl acetate. The organic phases were combined, washed with H₂O,dried (Na₂SO₄) and concentrated. The crude product was purified bychromatography (silica gel, CH₂Cl₂-MeOH gradient) to afford Example 10as a solid: C₂₆H₂₉N₇O₃: m/z MH⁺ 488; ¹H NMR (300 MHz, DMSO) δ 2.28 (s,6H), 2.51-2.63 (m, 2H), 3.80 (s, 3H), 4.14-4.44 (overlapping m, 5H),5.68-5.76 (m 1H), 6.11-6.19 (m, 1H), 6.43-6.48 (m, 1H), 6.95 (s, 1H),7.11-7.17 (m, 1H), 7.37-7.45 (overlapping m, 2H), 7.68-7.07 (d, 1H,J=8.4 Hz), 8.39-8.43 (overlapping m, 4H), 9.75 (s, 1H) ppm.

Example 11N-(2-(2-(Dimethylamino)ethoxy)-5-((4-(1-(2-fluoroethyl)-1H-indolyl-3-yl)pyrimidin-2-yl)amino)-4-methoxyphenyl)acrylamide(11)

1-(2-Fluoroethyl)-1H-indole (Scheme 9). Sodium hydride, 60 wt. % in oil(2.3 g, 57.5 mmol) was added to stirred, 0° C., clear, colorlesssolution of indole (10.1 g, 86.2 mmol) in anhydrous tetrahydrofuran atas rapid a rate consistent with maintaining control of the concomitanthydrogen evolution. Solution was stirred at 0° C. under N₂ blanket untilgas evolution ceased, and reaction had become a fine white suspension. Asolution of 1-fluoro-2-iodoethane (5 g, 29 mmol) in anhydroustetrahydrofuran (6 mL) was then slowly added via syringe, the ice bathwas removed and the pot heated to reflux overnight. The reaction mixturewas cooled, diluted with a solution of ammonium chloride (4.6 g, 86mmol) in DI water (300 mL), transferred to a separatory funnel, andextracted with ethyl acetate. The extract was dried (CaSO₄) andevaporated to provide a yellow oil, which was flash chromatographed(silica gel, 100% hexanes) to provide 4.2 g of yellow oil, characterizedby LC-MS as a 60/40 mixture of indole to desired product. This impureproduct was treated with benzene sulfonyl chloride to modify the elutioncharacteristics of the mixture to allow for isolation of the desiredproduct as follows: To a 0° C. solution of the above isolated 60/40mixture of indole to desired product and tetrabutyl ammonium bisulfate1.2 g, 3.4 mmol) in anhydrous toluene (100 mL) was added a solution ofsodium hydroxide (24.7 g, 617.5 mmol) in DI water (25 mL). To therapidly stirred, 0° C., mixture was then added benzene sulfonyl chloride(5.5 mL, 43.1 mmol) and the reaction allowed to stir and warm to ambienttemperature under N₂ blanket overnight.

The reaction mixture was then partitioned between ethyl acetate and DIwater, the organic phase dried (CaSO₄) and flash chromatographed (silicagel, 10% acetone/hexanes) to cleanly resolve the 1-phenylsulfonyl indolefrom the desired product affording 1.3 g of 1-(2-fluoroethyl)-1H-indoleas a clear, colorless liquid. ¹H NMR (300 MHz, DMSO) δ 4.45 (t, 1H,J=4.9 Hz), 4.54 (t, 1H, J=4.9 Hz), 4.64 (t, 1H, J=4.6 Hz), 4.80 (t, 1H,J=4.4 Hz), 6.46 (dd, 1H, J=3.1, 0.8 Hz), 7.03 (m, 1H), 7.13 (m, 1H),7.37 (d, 1H, J=3.2 Hz), 7.49 (d, 1H, J=8.3 Hz), 7.55 (m, 1H) ppm. ¹³CNMR (75 MHz, DMSO) δ 46.4 (d, J_(CF)=19.5 Hz), 83.3 (d, J_(CF)=166.5Hz), 101.4, 110.3, 119.6, 120.9, 121.6, 128.6, 129.3, 136.4 ppm.C₁₀H₁₀NF m/z MH⁺ 164.

3-(2-Chloropyrimidin-4-yl)-1-(2-fluoroethyl)-1H-indole (Scheme 9,Intermediate U). Ferric chloride (1.3 g, 7.9 mmol) was rapidly added toa stirring, ambient temperature, degassed, clear, colorless solution of1-(2-fluoroethyl)-1H-indole and 2,4-dichloropyrimidine (1.2 g, 8.3 mmol)dissolved in anhydrous 1,2-dimethoxyethane (80 mL). The resultant black,opaque, solution was stirred at 60° C. for 17 h under dry nitrogenatmosphere, cooled, and partitioned between ethyl acetate and saturatedaqueous sodium chloride. The organic phase was dried (CaSO₄) andevaporated to provide 2.3 g of purple oil which was purified by flashchromatography (silica gel, 0 to 90% ethyl acetate in hexanes) to yield557.5 mg of 3-(2-chloropyrimidin-4-yl)-1-(2-fluoroethyl)-1H-indole (U)as a light yellow powder. ¹H NMR (300 MHz, DMSO) δ 4.60 (t, 1H, J=4.7Hz), 4.69 (t, 1H, J=4.8 Hz), 4.75 (t, 1H, J=4.4 Hz), 4.90 (t, 1H, J=4.4Hz), 7.31 (m, 2H), 7.67 (m, 1H), 7.88 (d, 1H, J=5.5 Hz), 8.44 (m, 1H),8.57 (m, 2H) ppm. ¹³C NMR (75 MHz, DMSO) δ 47.2 (d, J_(CF)=19.8 Hz),82.8 (d, J_(CF)=167.7 Hz), 111.6, 111.9, 115.0, 122.1, 122.3, 123.4,125.8, 134.6, 137.8, 159.4, 160.8, 164.9 ppm. C₁₄H₁₁ClFN₃ m/z MH⁺ 276.

N-(4-Fluoro-2-methoxy-5-nitrophenyl)-4-(1-(2-fluoroethyl)-1H-indol-3-yl)pyrimidin-2-amine(Scheme 9, Intermediate V). p-Toluene sulfonic acid monohydrate (442.8mg, 2.3 mmol) was added to a stirred suspension of3-(2-chloropyrimidin-4-yl)-1-(2-fluoroethyl)-1H-indole (U) (535.3 mg,1.9 mmol) and 4-fluoro-2-methoxy-5-nitroaniline (361.4 mg, 1.9 mmol) in1,4-dioxane (20 mL) and heated to reflux under nitrogen blanket. Whileapproaching reflux temperature the suspended solid dissolved. Reflux wascontinued overnight, then the reaction was cooled and poured into arapidly stirred 5% (w/v) solution sodium hydrogen carbonate in DI water(200 mL) to precipitate product. Product was isolated by filtration,washed with water and allowed to dry to yield 921.4 mg ofN-(4-fluoro-2-methoxy-5-nitrophenyl)-4-(1-(2-fluoroethyl)-1H-indol-3-yl)pyrimidin-2-amine(V) as a fine yellow powder. C₂₁H₁₇F₂N₅O₃ m/z MH⁺=426.

N-(4-(2-(Dimethylamino)ethoxy)-2-methoxy-5-nitrophenyl)-4-(1-(2-fluoroethyl)-1H-indol-3-yl)pyrimidin-2-amine(Scheme 9, Intermediate W). 2-(Dimethylamino)ethanol (0.8 mL, 7.7 mmol)was slowly added to a stirred, N₂ blanketed, ambient temperature,suspension of sodium hydride, 60 wt. % in oil (306.4 mg, 7.7 mmol) inanhydrous 1,4-dioxane (24 mL). Anion formation was allowed to proceedfor 0.5 h, thenN-(4-fluoro-2-methoxy-5-nitrophenyl)-4-(1-(2-fluoroethyl)-1H-indol-3-yl)pyrimidin-2-amine(intermediate V) (652.0 mg, 1.53 mmol) was added all at once. Thereaction immediately turned to a red color, and was allowed to stir.After 10 min., LC-MS reported the reaction to be complete. DI water (5mL) was added to quench, then the mixture was partitioned between ethylacetate and saturated aqueous sodium chloride. The organic extract wasdried (CaSO₄) and evaporated to afford a yellow solid. This solid wasrecrystallized from boiling ethyl acetate/heptane, which upon cooling,precipitated a bright yellow crystalline powder. The powder was isolatedby filtration, washed with heptane, and allowed to dry providing 572.0mg ofN-(4-(2-(dimethylamino)ethoxy)-2-methoxy-5-nitrophenyl)-4-(1-(2-fluoroethyl)-1H-indol-3-yl)pyrimidin-2-amine(W). ¹H NMR (300 MHz, DMSO) δ 2.27 (s, 6H), 2.71 (t, 2H, J=5.7 Hz), 4.01(s, 3H), 4.33 (t, 2H, J=5.6 Hz), 4.56 (t, 1H, J=4.6 Hz), 4.65 (t, 1H,J=4.6 Hz), 4.73 (t, 1H, J=4.2 Hz), 4.89 (t, 1H, J=4.6 Hz), 7.01 (s, 1H),7.10 (m, 1H), 7.25 (m, 2H), 7.61 (d, 1H, J=8.4 Hz), 8.22 (s, 1H), 8.36(m, 3H), 8.76 (s, 1H) ppm. ¹³C NMR (75 MHz, DMSO) δ 46.2, 47.0 (d,J_(CF)=19.5 Hz), 57.3, 58.0, 69.0, 82.8 (d, J_(CF)=166.6 Hz), 99.2,108.2, 111.1, 113.4, 119.2, 121.4, 122.4, 122.6, 122.8, 126.0, 131.3,132.8, 137.6, 150.6, 156.2, 157.7, 160.5, 162.5 ppm. C₂₅H₂₇ F N₆O₄ m/zMH⁺=495.

4-(2-(Dimethylamino)ethoxy)-N1-(4-(1-(2-fluoroethyl)-1H-indol-3-yl)pyrimidin-2-yl)-6-methoxybenzene-1,3-diamine(Scheme 9, Intermediate X). Stannous chloride dihydrate (708.3 mg, 3.1mmol) was added to a stirred, ambient temperature yellow suspension ofN-(4-(2-(dimethylamino)ethoxy)-2-methoxy-5-nitrophenyl)-4-(1-(2-fluoroethyl)-1H-indol-3-yl)pyrimidin-2-amine(W) (303.8 mg, 0.6 mmol) in ethyl acetate (30 mL) and heated at refluxunder nitrogen blanket for 4 h. The reaction was allowed to cool, thenpoured into a 5% (w/v) solution of sodium hydrogen carbonate in DI water(200 mL) and stirred for 0.5 h. The multiphase mixture was then filteredthrough tightly packed celite, with ethyl acetate rinsing of the filtercake. The filtrate was transferred to a separatory funnel and the liquidphases separated. The retained ethyl acetate solution of product waswashed with saturated aqueous sodium chloride, dried (CaSO₄), andevaporated to provide a red oil which was purified by flashchromatography (silica gel, 2% NH₄OH(aq.) in methanol/ethyl acetate; 0to 10%) to isolate X as 165.4 mg of red oil. C₂₅H₂₉FN₆O₂ m/z MH⁺=465.

N-(2-(2-(Dimethylamino)ethoxy)-5-((4-(1-(2-fluoroethyl)-1H-indol-3-yl)pyrimidin-2-yl)amino)-4-methoxyphenyl)acrylamide(11, Scheme 9). 3-Chloropropanoyl chloride (38 mL, 0.4 mmol) was slowlyadded, by syringe, to a rapidly stirred, 0° C., nitrogen blanketedsolution of4-(2-(dimethylamino)ethoxy)-N1-(4-(1-(2-fluoroethyl)-1H-indol-3-yl)pyrimidin-2-yl)-6-methoxybenzene-1,3-diamine(Intermediate X) in anhydrous tetrahydrofuran (20 mL). Upon thisaddition, precipitate immediately formed. The suspension was stirred at0° C. for an additional 5 min. then the ice bath was removed. Uponconfirmation of complete conversion to the 3-chloropropanamideintermediate, a solution of sodium hydroxide (726.0 mg, 18.2 mmol) in DIwater (5.0 mL) was added to the reaction suspension which was heated toreflux for 1 h then cooled and partitioned with brine and additionaltetrahydrofuran. The organic extract was dried (CaSO₄) and evaporated toyield 445.1 mg of solid orange foam which was purified by gradient flashchromatography (silica gel, 2% NH₄OH(aq.) in methanol/ethyl acetate; 0to 10%), and crystalized from ethyl acetate/heptane to isolate 130 mg ofExample 11 as a fine light yellow powder. ¹H NMR (300 MHz, DMSO) δ 2.28(s, 6H), 2.58 (t, 2H, J=5.3 Hz), 3.86 (s, 3H), 4.19 (t, 2H, J=5.3 Hz),4.58 (t, 1H, J=4.6 Hz), 4.67 (t, 1H, J=4.5 Hz), 4.72 (t, 1H, J=4.6 Hz),4.88 (t, 1H, J=4.6 Hz), 5.75 (dd, 1H, J=10.4, 1.7 Hz), 6.22 (dd, 1H,J=17.0, 1.9 Hz), 6.48 (m, 1H), 6.95 (s, 1H), 7.14 (t, 1H, J=7.4 Hz),7.22 (m, 2H), 7.60 (d, 1H, J=8.2 Hz), 7.94 (s, 1H), 8.30 (m, 2H), 8.56(s, 1H), 8.80 (s, 1H), 9.83 (s, 1H) ppm. ¹³C NMR (75 MHz, DMSO) δ 45.6,46.9 (d, J_(CF)=19.9 Hz), 56.6, 57.9, 60.2, 69.4, 82.9 (d, J_(CF)=168.2Hz), 101.6, 107.5, 111.1, 113.6, 116.9, 121.4, 122.3, 122.6, 123.2,126.0, 126.6, 132.6, 133.2, 137.6, 145.3, 147.8, 158.0, 160.7, 162.1,163.2 ppm. C₂₈H₃₁FN₆O₃ m/z MH⁺=519.

The following non-limiting Examples further illustrate certain aspectsof the present invention, which are prepared according to the generalSynthetic Schemes 1 to 9 above:

Biological Assays

Compounds of the formula I as novel EGFR tyrosine kinase inhibitors wereevaluated for their activity against EGFR according to the proceduresdescribed below.

Cell Culture. A431 (passage 3) and NCI-H1975 (passage 5) cells (ATCC)were started from frozen stocks and cultured in RPMI 1640 mediumsupplemented with 10% heat-inactivated fetal bovine serum, 1×penicillin/streptomycin/glutamine, 1 mM sodium pyruvate, 10 mM4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES), and 0.25%D-glucose (growth medium) in T175 flasks in a humidified 30° C., 5% CO₂incubator. The cell monolayer was dispersed by 5 minute exposure to0.25% Trypsin/EDTA solution (Life Technologies) and the solution wasneutralized with a fresh growth medium. Pooled cells were pelleted bycentrifugation (200× g, 8 min.), resuspended in the growth medium, andan aliquot was removed for cell counting using an automated cell counter(Logos Biosystems). The cells maintained normal morphology and growthcharacteristics during the period of the study.

Cell Proliferation Assay. Dispersed cells were pooled by centrifugation(200×g, 8 min.) and resuspended in a fresh medium to a concentration of1.00 E+04 cells/ml. 200 μL of the cell suspension was added to each well(2,000 cells/well) of a black-walled 96 well plate and the cells wereallowed to attach overnight under normal culture conditions. Afterovernight culturing, 1 μL of a test compound (n=3 per concentration) wasadded per well to achieve final concentrations of 10, 3.33, 1.11, 0.370,0.124, 0.0412, 0.0137, 0.0046, and 0.0015 μM. The final DMSOconcentration in the well was 0.5% v/v. Vehicle, non-treated, andcell-free wells were also included in the assay. The cells were culturedunder normal conditions for 72 hours with daily visual inspection.

Cell proliferation was measured using the dye Alamar Blue (resazurin).Resazurin is reduced by cellular enzymes to resorufin, which isfluorescent (544 nM excitation, 612 nm emission). Fluorescence intensitywas proportional to cell number. A resazurin stock solution was preparedin a phosphate-buffered saline (PBS) to a stock concentration of 440 μM.The resazurin stock solution (40 μL each) was added to each well at hour67 of the 72 hour incubation period. The plate was returned to normalculture conditions and fluorescence measurements were collected using aCytation 3 multimode plate reader (Biotek) at 72 hours.

Data Analysis. Fluorescence measurements were normalized againstcell-free (background) readings and the total growth over 72 hour timeperiod was determined versus the average of the vehicle control wells.Average and standard deviation values were determined for each condition(n=3).

Table 1 contains illustrative data from study of representativecompounds of the present invention, which demonstrate their excellentselectivity for inhibition of the growth of H1975 (double mutant) cellsover A431 (wild type) cells.

TABLE 1 Biological activity of selected compounds in the A431 (wildtype) and H1975 (double mutant) cell proliferation assays. Example A431IC₅₀ (μM)^(a) H1975 IC₅₀ (μM)^(a) 1 + +++ 2 + +++ 3 + ++ 4 + +++ 5 ++ ++8 ++ +++ 9 + +++ 10 ++ +++ 11 ++ +++ ^(a)An IC₅₀ value greater than 1.0μM is represented by “+”; an IC₅₀ in the range of 0.1-1.0 μM isrepresented by “++”, and an IC₅₀ value below 0.1 μM is represented by“+++”.

The in vivo anticancer activity of Examples 1 and 2 is also illustratedin FIGS. 1-4.

Antitumor Activity of Example 1 in the H1975 Mouse Xenograft Model. Thein vivo anticancer activity of Example 1 against tumors with theL858R/T790M double mutation is illustrated in FIG. 1. Example 1 wasevaluated in subcutaneously-implanted H1975 human non-small cell lungcarcinoma xenographs in female nude mice at 6.25, 12.5 and 25 mg/kg.Example 1 was dosed orally once a day for 14 days (days 6-19). At alldoses, Example 1 was well tolerated, resulting in no treatment-relatedmortality. Treatment with 1 at 6.25, 12.5 and 25 mg/kg produced a mediantime to evaluation size of 28.9, 31.6 and 34.3 days, respectively,resulting in a statistically significant (P<0.05) tumor growth delay of14, 16.7 and 19.3 days, respectively. At 25 mg/kg, treatment produced a100% incidence of complete regressions and 10% of the mice were tumorfree survivors.

Antitumor Activity of Example 2 in the H1975 Mouse Xenograft Model. Thein vivo anticancer activity of Example 2 against tumors with theL858R/T790M double mutation is illustrated in FIG. 2. Example 2 wasevaluated in subcutaneously-implanted H1975 human non-small cell lungcarcinoma xenographs in female nude mice at 50 and 100 mg/kg. Example 2was dosed orally once a day for 14 days (days 7-20). At 100 mg/kg oraldosing, Example 2 was well tolerated and produced significant (P<0.05)anticancer activity based upon the % tumor growth inhibition values (%TGI) of 110.5%, 116.6% and 116.6%, which were calculated from the mediantumor burdens on days 10, 14 and 17, respectively. Time to evaluationsize (750 mm³) was 39.6 days, resulting in a tumor growth delay (T-C) of22.2 days, which is also statistically significant. Treatment produced a100% incidence of complete tumor regression and 12.5% of the miceremained tumor free (TFS) at the completion of the study.

Antitumor Activity of Example 1 in the HCC827 Mouse Xenograft Model. Thein vivo anticancer activity of Example 1 against tumors with thedelE746-A750 activating mutation is illustrated in FIG. 3. Example 1 wasevaluated in a subcutaneously-implanted HCC827 human non-small cell lungcarcinoma xenographs in female nude mice at 6.25 mg/kg. Example 1 wasdosed orally once a day for 14 days (days 13-26). At 6.25 mg/kg oraldosing, 1 was well tolerated, resulting in no treatment-relatedmortality. Treatment with 1 produced a median time to evaluation size of61.5 days, resulting in a statistically significant (P<0.05) tumorgrowth delay of 33.2 days. Treatment produced a 100% incidence ofcomplete tumor regression at the completion of dosing. FIG. 4 shows theaverage concentration of Example 1 in plasma, brain and tumor tissuesfollowing a 25 mg/kg oral dose in this model.

The foregoing examples and description of the preferred embodimentsshould be taken as illustrating, rather than as limiting the presentinvention as defined by the claims. As will be readily appreciated,numerous variations and combinations of the features set forth above canbe utilized without departing from the present invention as set forth inthe claims.

What is claimed is:
 1. A method of treating a disease or disorderassociated with mutant EGFR activity, comprising administering to apatient in need of treatment a therapeutically effective amount of acompound of formula 1, or a pharmaceutically acceptable salt thereof

wherein the disease or disorder is associated with a delE746-A750activating mutant of EGFR.
 2. The method of claim 1, wherein the diseaseor disorder is a cancer selected from the group consisting of braincancer, lung cancer, kidney cancer, bone cancer, liver cancer, bladdercancer, head and neck cancer, esophageal cancer, stomach cancer, coloncancer, rectum cancer, breast cancer, ovarian cancer, melanoma, skincancer, adrenal cancer, cervical cancer, lymphoma, and thyroid tumorsand their complications.
 3. The method of claim 2, wherein the cancer isbrain cancer or lung cancer.
 4. The method of claim 1, in combinationwith administering to the patient a second therapeutic agent.
 5. Themethod of claim 4, wherein the second therapeutic agent is achemotherapeutic agent.
 6. The method of claim 4, wherein the secondtherapeutic agent is a different EGFR modulator.
 7. A method ofpreparing a compound of formula (I), comprising reacting an intermediatecompound C with 3-chloropropionyl chloride in the presence of a base toform the compound of formula (I):

wherein G is 1-methyl-1H-indol-3-yl, 1-(2-fluoroethyl)-1H-indol-3-yl,1,2-dimethyl-1H-indol-3-yl, pyrazolo[1,5-a]-pyridin-3-yl, or1-methyl-1H-indazol-3-yl; and X is O, S, or CH₂ wherein: when X is O orS, further comprising converting compound A to intermediate compound Bthrough displacement of the fluoro (F) group in the compound A byHX(CH₂)₂N(CH₃)₂ in the presence of a base and converting theintermediate compound B to the intermediate compound C by reduction ofthe nitro group on the compound B:

when X is CH₂, further comprising converting compound A to intermediatecompound D by displacement of the fluoro (F) group in the compound Awith 3-dimethylamino-1-propynyl lithium and converting the compound D tothe compound C through hydrogenation of the compound D in the presenceof a palladium or platinum catalyst: